The pathogenicity of Corynebacterium diphtheriae, causing diphtheria in man, is primarily due to the production of a potent proteinaceous exotoxin, The genetic information for synthesis of this toxin resides in the DNA of certain corynebacteriophages. This system is an excellent model of how virus-infected pathogenic bacteria may release potent toxins which interfere with metabolism in man and cause diseases such as scarlet fever, botulism, staphylococcal enteritis and diphtheria. Diphtheria toxin inhibits cellular protein synthesis in susceptible animals including man by ADP-ribosylation of elongation factor 2. Maximum toxin synthesis by C. diphtheriae occurs at less than 75 microgram/l iron in the growth medium; higher iron concentrations are inhibitory. The objective of the proposed research is to determine if toxin synthesis is controlled by iron (1) at the level of transcription of toxin mRNA, (2) during translation of toxin mRNA, or (3) during release of toxin through the cell membrane. A cell-free protein synthesizing system from E. coli will be used to compare toxin synthesis by mRNAs extracted from cells growing in high and low iron concentrations. Extracts of C. diphtheriae grown at high and low levels of iron will be tested in this E. coli system for inhibitory effects on toxin mRNA translation. Five representative lysogenic, toxigenic strains will be examined and compared for cellular levels of toxin mRNA by means of DNA-RNA molecular hybridization using DNA of a toxigenic phage and these data will be compared to ability to synthesize and to excrete diphtheria toxin. Cell membrane lipids of representative strains growing in different concentrations of iron will be extracted and examined by gas-chromatography, mass-spectrometry for possible involvement of specific lipids in control of extracellular release of toxin.